Interleukin-2 stimulation induces tyrosine phosphorylation of p120-Cbl and CrkL and formation of multimolecular signaling complexes in T lymphocytes and natural killer cells.
Interleukin (IL)-2, a major growth and differentiation factor for T lymphocytes, was found to induce tyrosine phosphorylation of the proto-oncogene products p120-Cbl and CrkL in IL-2-dependent cell lines. We established that, in unstimulated lymphocytes, the Src homology 2 (SH2) and SH3 domain-containing protein Grb2 and the p85 subunit of phosphatidylinositol 3-kinase, associate constitutively with Cbl via their SH3 domains. Furthermore, IL-2 stimulation increased the level of interaction of phosphorylated Cbl with the p85 SH2 domains, and we provide evidence that the preformed Cbl-Grb2 complex recruits the phosphorylated p52 Shc adaptor protein. In addition, we demonstrate that the SH2-SH3-SH3 adaptor protein CrkL is tyrosine-phosphorylated in an IL-2-dependent manner and, via its SH2 domain, associates with a large proportion of phosphorylated Cbl. We also show that p85 is preassociated with the CrkL SH3 domain. Furthermore, the association of CrkL and p85 is increased after IL-2 treatment by a mechanism involving intermediary tyrosine-phosphorylated proteins that remain to be identified. Our results show that CrkL associates independently with Cbl or p85 and suggest that it also participates in larger complexes containing Cbl and p85. Although the precise roles of Cbl and CrkL remain to be elucidated, their tyrosine phosphorylation, in addition to the multiple protein interactions described here, strongly suggest that Cbl and CrkL may play pivotal roles in the early steps of IL-2 signal transduction.
Gesbert
F
F
INSERM Unit 461, Faculté de Pharmacie Paris-XI, 92296 Châtenay-Malabry, France.
Garbay
C
C
Bertoglio
J
J
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology*; D007694:Killer Cells, Natural / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000032:analysis; D011506:Proteins / Q000378:metabolism; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D050721:Proto-Oncogene Proteins c-cbl; D015398:Signal Transduction / Q000502:physiology; D013601:T-Lymphocytes / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D018909:src Homology Domains / Q000502:physiology
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; D007376:Interleukin-2; D009687:Nuclear Proteins; D019869:Phosphatidylinositol 3-Kinases; D019000:Phosphotyrosine; D011506:Proteins; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins; D050721:Proto-Oncogene Proteins c-cbl; D014443:Tyrosine; D044767:Ubiquitin-Protein Ligases; C082869:CRKL protein; C495948:GRB2 protein, human; C000715470:MAS1 protein, human; C067419:CBL protein, human